D‐2‐hydroxyglutarate suppresses allergic sensitization in a murine model of experimental asthma
Authors: Anuj Tharakan, Ankit Kumar, Jeremy Allegood, Lauren Ashley Cowart, Daniel H. Conrad, Rebecca K. Martin
Journal: Allergy (2023)

Asthma is a chronic inflammatory disease affecting the airways which is characterized by reversible airflow obstruction, bronchial hyperresponsiveness, and airway remodeling. In allergic asthma, this airway pathology is mediated by Type 2 cytokines (IL-4, IL-5, and IL-13) and allergen-specific IgE. IL-4, IL-5, and IL-13 are predominantly produced by lung resident T helper 2 (Th2) cells and Type 2 innate lymphoid cells. The production of allergen-specific IgE from B cells is driven by a subset of IL-4 and IL-13 producing T follicular helper cells termed T follicular helper 13 (Tfh13) cells.1, 2 The initiation of these T-cell responses is mediated by dendritic cells (DCs). The mechanisms by which DCs regulate Th2 and Tfh13 polarization, however, are unclear. Several genome-wide association studies have identified that single-nucleotide polymorphisms (SNPs) in the D2HGDH locus are associated with allergic asthma and allergic rhinitis risk, with the G allele of rs34290285 associated with a lower risk of disease.3 The D2HGDH gene encodes the enzyme D-2-hydroxyglutarate dehydrogenase (D2HGDH), which converts D-2-hydroxyglutarate (D2HG) to α-ketoglutarate (α-KG). To elucidate the functional consequences of these SNPs, the regulatory landscape of the D2HGDH locus was analyzed using the ENCODE database. We found that the sentinel SNP, rs34290285, resides within a candidate cis-regulatory element (cCRE) with distal enhancer-like features in human monocytes (Figure 1A). Previously described lung eQTL analysis of the 10 most significant SNPs revealed that these asthma risk alleles are associated with reduced expression of D2HGDH mRNA in the lung (Figure 1B).3 As the lead SNP, rs34290285 is also associated with GAL3ST2, the effects of GAL3ST2 cannot be excluded.3 The downregulation of D2HGDH expression suggests that these SNPs may promote accumulation of D2HG, which is a potent endogenous inhibitor of α-KG-dependent dioxygenases, a group of histone demethylase and prolyl hydroxylase enzymes.4 Recent studies demonstrate that inhibition of these α-KG dependent histone demethylases in DCs results in a tolerogenic DC phenotype.5 D-2HG and α-KG, therefore, reciprocally regulate these dioxygenase enzymes to dynamically regulate gene expression via epigenetic mechanisms. Thus, we performed metabolomic analysis of murine bone marrow-derived dendritic cells following stimulation with LPS or the allergen Alternaria which revealed a significant increase in the ratio of α-KG to hydroxyglutarate following Alternaria treatment (Figure 1C). Therefore, we predicted that D-2HG may modulate DC function to alter allergic sensitization in a murine model of allergic airway inflammation. We found that intranasal administration of octyl-D-2HG, a cell permeable form of D-2HG, prior to intranasal exposure to Alternaria significantly reduced mediastinal lymph node (mLN) Th2, Tfh13, and Tfh2 cell polarization, but overall Tfh priming was unimpaired (Figure 2A–D. Figure S1A,B). Further, octyl-D2HG impaired mLN IgE+ germinal center B cell (GCBC) and IgE+ plasma cell (PC) differentiation (Figure 2E,F). IgG1+ GCBC and PC differentiation, however, was unaffected by octyl-D2HG administration (Figure S1C,D). Additionally, octyl-D-2HG administration promoted T follicular regulatory (Tfr) cell polarization, which act to restrain IgE production in allergic asthma (Figure 2G).6 Intranasal treatment of octyl-D2HG in a model of established murine allergic asthma resulted in a durable suppression of airway hyperreactivity for 4 days following treatment (Figure 2H,I) Additionally, mice treated with octyl-D2HG exhibited reduced levels of airway antigen-specific IgE and MCPT-1, indicating suppression of airway mast cell degranulation (Figure 2J,K). Overall, these data demonstrate that D2HG critically regulates allergic sensitization and the polarization of Th2 and Tfh13 cells following allergen exposure. Further, we demonstrate that D-2HG induces Tfr cell differentiation in response to allergens in a murine model of allergic asthma, but it is not clear if this is the mechanism by which D2HG reduces allergic airway inflammation. Octyl-D2HG treatment in allergen-sensitized mice demonstrated significant attenuation of airway hyperreactivity and may therefore be a potential therapeutic approach for the treatment of allergic asthma and other IgE-mediated diseases. A.T. and R.K.M. made substantial contributions to concept and design. A.T., A.K., J.A., and R.K.M. made substantial contributions to analysis and/or interpretation of data. All authors were involved in drafting and/or critically revising the manuscript. None. This work was supported by funding from VCU's CTSA (UL1TR002649 from the National Center for Advancing Translational Sciences) to R.K.M. and the CCTR Endowment Fund of VCU. Services and products in support of the research project were generated by the Virginia Commonwealth University (VCU) Massey Cancer Center Flow Cytometry Shared Resource and Metabolomics and Lipidomics Shared Resource supported, in part, with funding from NIH-NCI Cancer Support Grant P30 CA016059. Rebecca K. Martin and Anuj Tharakan own stock in Pleros Therapeutics Inc. which is currently of no value. The rest of the authors have no conflicts of interest to declare. The data that support the findings of this study are available from the corresponding author upon reasonable request. Data S1 Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.